Entering edit mode
5.4 years ago
anjuraas
▴
10
hi I aligned my sequence with bowtie2 and put the BAM file into featurecount. And later the count table into deseq, but it is showing 'all samples have 0 counts for all genes'. Why my samples don't have miRNA?i downloaded the dataset from SRA database. please help
can you post a
headof your count table?and your
featureCountcommandI am using usegalaxy.
did you get any runtime log from it? are you using the correct gff (annotation) files?
Please confirm that the chromosome identifiers match between your gff and alignment.
yes. i checked that. Gene identifier ID is 'ID'. But its still showing 0 counts. Is there a chance that my read doesn't have miRNA?
What WouterDeCoster means is : are the same IDs present in all your files? eg. in GFF file it says
chr0and in your mappingChr00or such?Can you post a sample of the GFF you are using and of the genomic sequence (mainly the headers) you're mapping against?
Indeed, and more commonly, for human genetics, for example GFF:
chr1, alignment/fasta:1.This is my bowtie sam file
So your GFF contains NC_000001.11 and your sam file contains chr17... QED.
what can I do?please help
Use another gff which matches to the fasta you used for the alignment.
hi, I used the in built gff file,and now its showing few counts.but still I cant get the differential expression.this is the error report I got from deseq.i am sorry, I cant understand its meaning.
![Deseq error report][1]
Which organism are you working on? How was the data generated? Did you get appropriate alignment metrics?
I am working on dataset on gastric cancer downloaded from SRA database.its trueseq small RNA library using illumine Miseq. I aligned with bowtie 2 and got alignment rate above 90%.